Advanced age is the main risk factor for a range of human diseases. However, the fundamental mechanisms that drive aging remain largely unknown, impeding the development of interventions that delay or prevent age-related disorders and maximize the number of years without serious or chronic illnesses, referred to as healthspan. Senescent cells are known to accumulate in various tissues and organs with age and are thought to disrupt tissue structure and function by components they secrete. However, definitive evidence that senescent cells are causally implicated in aging and age-related disease has been lacking. To address the role of in vivo senescence in aging, we have generated a transgenic mouse model, termed INK-ATTAC, designed to inducibly kill pi 6'''''^-positive senescent cells upon administration of a synthetic drug. Our preliminary data indicate that the /A/K-ATTAC transgene inducibly kills pi 6''*^-positive senescent cells in the BubRI hypomorphic mouse model of premature aging, thereby delaying the onset of age-related pathologies. Based on these data we formulated the central hypothesis that cellular senescence is causally implicated in aging and that removal of senescent cells can prevent or delay age-related diseases and extend lifespan. We propose to test this hypothesis by pursuing three specific aims. In aim 1, we will identify p16'''^-positive senescent cells in mouse tissues and optimize conditions for /A//<-A7TAC-mediated killing. In aim 2, we will employ INK-ATTACXo prevent accumulation of p16''**-positive senescent cells throughout life and measure its impact on healthspan and lifespan. In aim 3, we will induce //VK-ATTAC-mediated clearance of senescent cells from aged mice to determine whether this treatment can delay or revert age-related diseases and extend lifespan. Based on our preliminary data in the premature aging model, we expect this subproject to reveal that senescent cells contribute to age-related loss of function and disease, and that elimination of senescent cells from tissues and organs increases healthspan in the absence of overt negative side effects. The research in this application is innovative, in our opinion, because it focuses on an entirely novel approach, senescent cell clearance, as a means to counteract aging-related functional decline and prevent chronic diseases and long-term illnesses. Overall, successful completion of Subproject 1 will significantly improve our understanding of the basic mechanisms of in vivo senescence and aging, and will lay the foundation for development of drugs that slow the development of aging-associated disorders by eliminating senescent cells, thus improving healthspan and quality of life of the elderly.